Project Summary/Abstract: Rett Syndrome (RTT) is a severe neurodevelopmental disorder characterized by a progressive loss of speech and social engagement, seizures, stereotyped hand movements, and motor system disabilities. The majority of RTT cases are caused by mutations in the methyl-DNA-binding protein MeCP2, but how mutations in MeCP2 lead to RTT symptoms is not fully understood. Our laboratory recently discovered that the loss of MeCP2 function in the brain in both RTT patients and MeCP2 null mice leads to a small but statistically significant and reproducible upregulation of genes that are long (>100kb), have high gene body levels of methyl-CA, and are enriched for neuronal function. However, the mechanism underlying long gene regulation by MeCP2 remains unknown, and it is unclear whether long gene upregulation is the cause of RTT symptoms. This proposal aims to answer these questions using a knock-in mouse model of RTT generated in our laboratory with one of the most common RTT patient mutations, MeCP2 R306C. MeCP2 R306C knock-in mice display behavioral features of RTT, upregulation of long, methyl-CA-enriched genes, and a loss of the MeCP2 interaction with the NCOR co-repressor complex. This suggests that the loss of the MeCP2-NCOR interaction in long genes could lead to long gene upregulation in RTT, and that long gene upregulation could cause RTT symptoms. The proposed study will test these hypotheses through the following aims: (1) to identify the genomic locations of altered NCOR complex binding, histone acetylation, and transcriptional elongation in MeCP2 R306C compared to WT mice, and (2) to characterize the synaptic defects in MeCP2 R306C mice and determine whether synaptic function can be rescued by reversing long gene upregulation. Understanding how the MeCP2 R306C mutation causes long gene upregulation and RTT symptoms will provide insight into the underlying cause of RTT, provide potential therapeutics for RTT, and contribute to an understanding of neuronal gene regulation that extends to other neurodevelopmental disorders.